There are an estimated 350 million people worldwide who are chronically infected with hepatitis B virus (HBV) and are at high risk for the development of hepatitis, cirrhosis (end stage liver disease) and hepatocellular carcinoma (HCC). Hepatitis and cirrhosis are associated with prolonged morbidity, and HCC, which appears in more than 250,000 people each year, is rapidly fatal. Given that the major risk factors for HCC are the HBV carrier state (defined as the persistence of virus or virus antigens in blood for more than 6 months) and chronic liver disease (CLD) (hepatitis and cirrhosis), the most direct way to reduce risk factors for HCC is to target the virus and/or corresponding immunity. Accordingly, the FDA has now approved interferon 1, pegylated interferon 1, lamivudine, adefovir dipivoxal, entecavir, and telbivudine. Additional drugs, such as emtricitabine, clevudine, and tenofovir, will probably be approved soon. Although these are potent drugs, they are limited by low rates of sustained response, side effects, and the emergence of drug resistance. Hence, the objective of this proposal is to devise combination therapies, following the model of HIV combination therapy development, to successfully treat long term HBV infection. To do so, the lab will assess the pharmacokinetics (PK), antiviral efficacy and toxicity of drug combinations in primary human hepatocytes and in HepG2 cells replicating wild type or selected mutants of HBV (aim 1). Favorable combinations will then be assessed in vivo in nude mouse injected with HepAD38 cells (AD38 cells replicate wild type HBV, which accumulates in the blood of these mice) in order to get a sense of what combinations and concentrations of drugs are most effective in reducing virus titer, for assessing how long virus DNA levels remain suppressed during and after treatment, and whether different combinations have a favorable PK profile in vivo (aim 2). The best combinations will then be evaluated for long term antiviral efficacy in HBV transgenic SCID mice, which stably replicate wild type HBV and develop CLD (aim 3). This work will develop combination therapies that will demonstrate a sustained antiviral effect against HBV, against HBV in HIV co-infected patients, in chronically infected alcoholics, as well as prolonged suppression of CLD. Elimination of CLD will reduce the risk of chronic HBV carriers of developing HCC. This will contribute centrally to the design of future clinical trials that will be suitable for long term treatment of chronic infections without significant side effects or the frequent development of drug resistance that limit present day therapeutics against HBV. PUBLIC HEALTH RELEVANCE There are more than 350 million people worldwide chronically infected with hepatitis B virus (HBV) and who are at risk for the development of hepatitis (inflammation of the liver), cirrhosis, and liver cancer. Millions of people each year die from liver cirrhosis. Liver cancer is among the top five most prevalent tumor types worldwide, with an estimated 1 million deaths each year. Present treatments for chronic hepatitis B use single drugs that are expensive, have side effects, and select for drug resistant viruses. Hence, new therapeutic options to treat long term virus infection and associated diseases are urgently needed. The proposed work will develop combination therapies with sustained antiviral effect against HBV and associated chronic liver disease, against HBV in HIV co-infected patients, and against HBV among chronic alcoholics. Prevention and/or elimination of chronic liver disease will reduce the risk of HBV carriers developing cirrhosis and liver cancer, thereby contributing centrally to improving public health.